Skeletal muscle differentiation is a well-orchestrated process regulated by autocrine, paracrine, and endocrine factors via a regulatory network of signal transduction pathways. In recent years the mammalian target of rapamycin (mTOR) has begun to be recognized as a critical regulator of skeletal muscle differentiation, growth and hypertrophy. Work from our laboratory has contributed to the current understanding of mTOR regulation of myoblast differentiation, and has led to the revelation that mTOR regulates multiple stages of myogenesis by assembling distinct pathways, some of which unexpected and yet to be fully delineated. With a combination of biochemical, molecular, cellular and genetic approaches, and utilizing both in vitro and in vivo systems, we aim to fill a sizable gap in the current knowledge of molecular pathways underlying the regulation of skeletal myogenesis by addressing these three major questions: (1) How are the known components of growth-regulating mTOR pathway involved in myogenesis, and what is the mTOR pathway(s) that regulates the initiation of myoblast differentiation in response to amino acids availability signals? (2) What is the mTOR pathway that specifically regulates the second-stage myocyte fusion critical for myotube/myofiber growth and maturation, and which secreted factors regulate this process? (3) What is mTOR's role in muscle regeneration and what are the mechanisms? Our expertise in biochemical characterization of signal transduction mechanisms, our strong preliminary data, and the unique animal models we have created, put us in an ideal position to tackle those questions. Knowledge gained in these studies will contribute to the molecular understanding of skeletal muscle development, repair, regeneration and hypertrophy. PUBLIC HEALTH RELEVANCE: Skeletal muscle differentiation is a well-orchestrated process regulated by autocrine, paracrine, and endocrine factors via multiple signal transduction pathways. Our proposed studies aim to dissect the molecular mechanisms underlying the regulation of skeletal muscle differentiation and regeneration, with a focus on the mammalian target of rapamycin signaling network. Knowledge gained in these studies will contribute to the molecular understanding of skeletal muscle biology, which may have significant impact on health-related issues such as muscular dystrophy, aging or disease-induced muscle atrophy, muscle regeneration, and exercise- induced muscle hypertrophy.